I. Technical Field
The present invention relates to a device for the aeration of liquid media by means of tube aeration, special tube modules contained therein and the use of a device for the aeration of liquid media.
II. Description of the Related Art
In the pharmaceuticals industry, the production of recombinant proteins is becoming ever more important. On account of its capabilities in producing glycosylated proteins with posttranslational modifications, animal cell culture has become well-established for the production of more complex proteins.
Usually intensive bubble aeration with dispersing stirring elements is used for supplying oxygen to microorganisms.
Animal cells impose special requirements on the technical reactors in which they are cultivated. High shearing forces, as are necessary for dividing up gas bubbles, are to be avoided in particular, since they irreversibly damage the cell membranes of the cells without cell walls and consequently have disadvantageous consequences for the growth of the cell cultures.
Apart from during dispersion by the stirring element, shearing forces are also released during the formation and bursting of the gas bubbles on the surface of the liquid. Bubble aeration can therefore only take place in cell cultivation with very low throughputs of gas. The stiffing element in this case does not undertake any gas bubble dispersing function but essentially only a distributing function. Therefore, in the case of cells that are sensitive to shearing, an adequate supply of oxygen can only be ensured to relatively low cell densities by an aeration method that uses large bubbles. (H. J. Henzler: “Verfahrenstechnische Auslegungsunterlagen für Rührbehäter als Fermenter” [process engineering design documents for stirring tanks as fermenters) Chem. Ing. Tech. 54 (1982) No. 5 pages 461-476, H. J. Henzler, J. Kauling: “Oxygenation of cell cultures” Bioprocess Engineering 9 (1993) pages 61-75, “Mischen and Rühren” [mixing and stirring] by M. Kraume, WILEY-VCH 2003)
To improve the oxygen supply, the use of sintered aerators of metallic or ceramic materials with pore widths of up to 0.2 μm has been proposed, allowing small bubbles to be produced with little shearing. (D. Nehring, P. Czermak, J. Vorlop, H. Lübben: “Experimental study of a ceramic micro sparging aeration system in a pilot scale animal cell culture” Biotechnology Progress 20 (2004)6, pages 1710-1717, Hanshi, Qi, Konstantin Konstantiniov: “The Art & Science of Micro-Sparging in High-Density Perfusion Cultures of Animal Cells” 17th ESTAC meeting, Tylosand, Sweden, 2001). However, when used over a long time, the sintered aerators have a tendency to become clogged. Furthermore, foaming problems may occur, requiring the use of anti-foaming agents, which in turn may lead to losses in yield in the reprocessing.
A low-shear method of supplying oxygen is represented by membrane aeration, in which the oxygen passes a membrane wall stretched between the gas phase and the culture medium. Such membranes can be wound up as tubes on cylindrical cage stators. (H. J. Henzler, J. Kauling: “Oxygenation of cell cultures” Bioprocess Engineering 9 (1993) pages 61-75, EP A1 0172478, WO A1 87/02054). To accommodate large exchange areas, the tubes are placed close together with as little spacing as possible.
The spacing S between the tubes, referred to the outside diameter of the tube D, is generally in the range of 0<S/D<1 in the case of suspended cells. If cells immobilized on carrier materials are used, greater spacings may also be advisable to ensure the permeability of the tube matrix with respect to the carrier particles. Both vertically and horizontally wound stators are used. Vertical winding is to be preferred in principle to prevent deposits on the tubes. Furthermore, in this way virtually double the tube area can be placed in the reactor. With the aid of low-shear radially transporting stirring elements such as blade or anchor stirrers, the concentrically arranged tube membranes are flowed through in the radial direction, in order to minimize the mass transfer resistance on the liquid side.
Other reactors, known as hollow-fiber reactors, connected in circular flow to a stirred tank reactor, are flowed through by medium (DE 195 37 033 A1, EP 0727 481 A2, DE 195 28 871 C2, U.S. Pat. No. 6,001,585 A, U.S. Pat. No. 5,443,985 A, WO 02/31108 A1). Here it is necessary for the culture solution to be constantly pumped around between the stirred tank reactor, in which the supply of media and the pH adjustment take place, and the aeration reactor. To be able to realize a high mass transfer, the mass transfer resistance on the liquid side at the membranes must be reduced, which requires high pumping circulation rates and consequently causes shearing problems. A further variant is the reciprocating jet reactor, in which the membrane areas are moved in an oscillating manner in the vertical direction. (Janine T. Bohlmann: “Entwicklung eines neuartigen Staustrahl-Membranreaktors für die Zellkulturtechnik mittels CFD” [development of a novel ramjet membrane reactor for cell culture technology by means of CFD] CIT, No. 75 (2003) pages 131-135). With such a configuration on an industrial scale, problems in handling the complicated membrane stator are likely, as well as technical problems in providing sterile conditions due to the sealing of the axial bearings. For the reasons stated, wound tube stators subjected to the flow of a coaxially arranged stirrer still appear to be the most technically sophisticated and reliable solution.
Silicone has been widely adopted in favor of porous polymers as the tube material. Reasons for this are the high gas permeability, the high thermal resistance and the tube properties distributed homogeneously over the length of the tube segments of over 50 m, properties that are even retained after sterilization. The great tube lengths of the tube segments serve for shortening the time-intensive production of the coaxial tube stators. The silicone tube is generally discarded after it has been used once, so that the considerable effort involved in producing an aeration cage is required each time after fermentation. The advantage of a less labor-intensive solution achieved by the use of long tube segments is also offset by disadvantages, however. For instance, compliance with an upper pressure loss limit for fixing the tube to distribution systems requires the use of large tube diameters in order to limit the pressure drop in the tube segments that is proportionately dependent on the ratio of tube length to tube diameter. Larger tube diameters in turn lead to a strength-dependent increase in the wall thickness, which causes a proportionate increase in the diffusion resistance in the mass transfer through the tube. A further considerable problem is represented by the reproducibility of the tube winding, which depends on the expenditure of force in stretching the tube, which cannot be definitively determined. In the case of vertical winding, a winding that is too tight leads to a cross-sectional restriction at the points of deflection and to a change in the pressure profile over the length of the tube. With tubes that are insufficiently stretched, there is the risk of the tubes being damaged by contact with the stirring element. A fundamental problem of membrane aeration is represented by the scaling of the specific tube surface area A/V, which decreases with the reactor volume V1/3, putting an upper limit on the scale of fermentation. It is not possible to compensate for this disadvantage by changed hydrodynamic conditions in the fermenter, for example by subjecting the tubes to improved flow with increased circumferential stirrer velocity, because of the increasing shear loading and the additional diffusion resistance, which cannot be hydrodynamically influenced.